Abstract

Glioblastoma multiforme (GBM) is the most lethal human brain tumor with high recurrence rates despite the existence of multimodal treatments including chemotherapy and radiation after surgical tumor resection. Augmented tumor metastasis after irradiation (IR) has been attributed to enhanced extracellular protease levels in tumors. Secreted or cleaved matrix metalloproteinase-2 (MMP-2) levels were reported to be elevated in irradiated tumors. Interestingly, previous in silico prediction analyses identified the presence of a putative nuclear localization signal (NLS) in MMP-2 and reported the cleavage of PARP by nuclear-MMP-2. However, MMP-2 nuclear localization in possible correlation with tumor progression has not been identified to date. In addition to membrane and cytoplasmic expression of MMP-2, we observed prominent MMP-2 nuclear localization, which is further elevated in IR (8 Gy)-treated human glioma xenograft cells 4910 and 5310. Interestingly, The protein-transcription factor (TF) interaction arrays using 4910 nuclear extracts revealed the potential binding of nuclear-MMP-2 with several transcription factors including CEF-2, c-Rel, FOXO4, GAS, GATA-1, MUSF-1, p300, PARP and RREB1. Co-IP experiments showed the direct binding between MMP-2/GATA-1 in glioma. Further, we also observed the presence of MMP-2 in the GATA-1 DNA-binding complex and its recruitment onto the promoter of IL-10, a GATA-1 target gene. ChIP-cloning with anti-MMP-2 antibody confirmed the specific binding of MMP-2 with GATA-1 consensus elements. We also observed that the MMP-2/GATA-1 nuclear co-localization was elevated in correlation with increasing GBM pathological grades. Downregulation of MMP-2 inhibited the GATA-1 expression and DNA-binding activity and abrogated IR-induced migration and invasion in these cells. Conversely, treatment with recombinant MMP-2 protein elevated the GATA-1 DNA-binding activity and increased IL-10 and phospho-Stat3 expression in correlation with increased invasion and migration. In vivo orthotopic experiments with MMP-2.siRNA decreased tumor growth in nude mice and inhibited nuclear MMP-2 and GATA-1 expression levels in tumors. To our knowledge, this is the first study demonstrating significant IR-induced nuclear translocation of MMP-2 and shows the functional association between MMP-2/GATA-1 in the regulation of IL-10/Stat3-mediated migration and invasion. Our results suggest the potential therapeutic efficacy of targeting MMP-2/GATA-1 liaison in glioma treatment.